---
res:
  bibo_abstract:
  - "This thesis comprises two distinct projects, each offering unique insights into
    fundamental\r\ncellular processes. While distinct in their focus, these different
    perspectives have a common\r\ntheme: chemiosmotic theory and utilisation of the
    proton gradient for driving the essential\r\nprocesses like auxin efflux and ATP
    synthesis, effectively bridging the membrane protein\r\nstructure and function
    from the realms of plant biology and cellular bioenergetics.\r\nThe first project
    of this thesis centres on the characterisation of PIN proteins, a class of\r\ntransmembrane
    transporters pivotal in the regulation of auxin transport and distribution in\r\nplants.
    PINs form a conserved and phylogenetically abundant group of transporters present
    in\r\nland plants and certain algae. Despite their great importance, they were
    one of the few elusive\r\nproteins essential for plant development not to be structurally
    and mechanistically\r\ncharacterised since their discovery almost 30 years ago.
    This work aimed to uncover the\r\nstructural and functional dynamics of the PIN
    protein-mediated auxin transport using an array\r\nof experimental techniques,
    including protein purification, biochemical assays and structural\r\nanalysis.
    Through an exhaustive screening process that took several years and included testing\r\ndifferent
    PIN homologues, expression systems, constructs, and purification conditions, we\r\ndeveloped
    a robust protocol for isolating the pure, stable, and monodisperse PIN8 protein.\r\nMoreover,
    utilising biophysical methods and buffer screening, we demonstrated that PIN8\r\nexhibits
    detergent and pH-dependent stability, with mild detergents and lower pH (5.0 and
    6.0)\r\nbeing optimal for the stability of the protein. Using SEC-MALS and crosslinking,
    we\r\ndetermined that PIN8 forms dimers, which was confirmed by our structural
    studies. We\r\nobtained a cryo-EM map of PIN8 at pH 6.0, and, compared to recently
    published structures,\r\nour map implies major pH-dependent conformational changes
    and possibly utilisation of the\r\nproton gradient in the transport mechanism.\r\nThe
    subject of the second project was F1Fo-ATP synthase, an enzyme complex fundamental\r\nto
    cellular energy metabolism. Through an approach integrating biochemical assays
    and\r\nstructural analysis, this research aimed to unveil the molecular mechanism
    of inhibition of ATP\r\nsynthase by yaku´amide, a bioactive compound with potential
    therapeutic implications. Using\r\nsubmitochondrial particles and purified F1Fo-ATP
    synthase, we demonstrated that, contrary to\r\npublished data, yaku´amide inhibits
    both ATP hydrolysis and ATP synthesis reactions.\r\nMoreover, we found that yaku´amide
    inhibitory activity is proton motive force (pmf)\r\ndependent, with lower inhibition
    in a more coupled system. Utilising cryo-EM, we obtained\r\nmaps and models for
    the three main rotational states of murine ATP synthase (State 1 at 3.0 Å,\r\n8\r\nState
    2 at 3.1 Å, and State 3 at 3.2 Å, overall). We observed several new features in
    our maps;\r\nhowever, we cannot definitively determine the exact mechanism of
    yaku amide’s inhibition on\r\nthe protein due to either resolution limits or suboptimal
    binding of the inhibitor.@eng"
  bibo_authorlist:
  - foaf_Person:
      foaf_givenName: Kristina
      foaf_name: Lukic, Kristina
      foaf_surname: Lukic
      foaf_workInfoHomepage: http://www.librecat.org/personId=2B04DB84-F248-11E8-B48F-1D18A9856A87
    orcid: 0000-0003-1581-881X
  bibo_doi: 10.15479/at:ista:17319
  dct_date: 2024^xs_gYear
  dct_isPartOf:
  - http://id.crossref.org/issn/2663-337X
  dct_language: eng
  dct_publisher: Institute of Science and Technology Austria@
  dct_title: 'Membrane proteins in plant physiology and bioenergetics : Investigating
    auxin efflux transporter PIN8 and ATP synthase inhibition by the novel inhibitor
    Yaku''amide B@'
...